Granulomatous enteritis in rainbow trout (Oncorhynchus mykiss) associated with soya bean meal regardless of water dissolved oxygen level.

This study investigated morphological changes associated with soya bean meal-induced enteritis (SBMIE) in distal intestine (DI) of rainbow trout (Oncorhynchus mykiss) fed a soya bean meal (SBM)-based diet and exposed to normoxia or hypoxia created by optimal and low water flow rates, respectively. A 28-day adaption period was followed by a 42-day challenge period where 600 fish were subjected to dietary challenge and/or hypoxia. Twelve tanks each containing 50 juvenile trout were assigned randomly in triplicate to each treatment. Histopathological and immunohistochemical evaluation revealed pathological features that have not previously been described in association with SBMIE. Vacuolar degeneration of epithelial cells mainly at the base of mucosal folds, epithelial cysts, epithelial dysplasia, necrosis, shedding of necrotic cells, and granulomatous inflammation including infiltration of enlarged, sometimes finely vacuolated or "foamy" macrophages, multinucleated giant cells and increased proliferation of fibroblasts were observed. Acid-fast bacteria were not detected in enlarged macrophages; however, these cells contained AB-PAS- and sometimes cytokeratin-positive material, which was interpreted to be of epithelial/goblet cell origin. Hypoxia did not affect the morphological changes in DI. These results suggest that SBM was associated with a granulomatous form of enteritis in DI of rainbow trout regardless of water oxygen level.

Loop-mediated isothermal amplification for detection of the tomato and potato late blight pathogen, Phytophthora infestans.

AIMS: To design and validate a colorimetric loop-mediated isothermal amplification assay for rapid detection of Phytophthora infestans DNA. METHODS AND RESULTS: Two sets of loop-mediated isothermal amplification (LAMP) primers were designed and evaluated for their sensitivity and specificity for P. infestans. ITSII primers targeted a portion of the internal transcribed spacer region of ribosomal DNA. These primers had a limit of detection of 2 pg P. infestans DNA and cross-reacted with the closely related species Phytophthora nicotianae. Rgn86_2 primers, designed to improve assay specificity, targeted a portion of a conserved hypothetical protein. These primers had a limit of detection of 200 pg P. infestans DNA and did not cross-react with P. nicotianae. The specificity of the Rgn86_2 assay was tested further using the closely related species P. andina, P. ipomoeae, P. mirabilis and P. phaseoli. Cross-reactions occurred with P. andina and P. mirabilis, but neither species occurs on tomato or potato. Both primer sets were able to detect P. infestans DNA extracted from tomato late blight leaf lesions. CONCLUSIONS: Two colorimetric LAMP assays detected P. infestans DNA from pure cultures as well as infected leaf tissue. The ITSII primers had higher sensitivity, and the Rgn86_2 primers had higher specificity. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report of a LAMP assay for the detection of P. infestans, the causal organism of potato and tomato late blight. These assays have potential for immediate utility in plant disease research and diagnostic laboratories.

First Report of the European Lineage of Phytophthora ramorum on Viburnum and Osmanthus spp. in a California Nursery.

Phytophthora ramorum S. Werres & A.W.A.M. de Cock is the causal agent of sudden oak death in California and Oregon forests and ramorum blight on a broad range of host species in wildlands and nurseries. It is thought to be an introduced pathogen and only three clonal lineages are known (3). The North American lineage (lineage NA1, mating type A2) is responsible for infections in California and Oregon forests. The European lineage (lineage EU1, predominantly A1) is responsible for infections in Europe, but has also been found in nurseries in Oregon and Washington. A third lineage (NA2) has only been isolated in a few instances from nurseries in Washington and California. In June 2006, P. ramorum was isolated from diseased Viburnum tinus, Osmanthus heterophyllus, and O. fragrans cultivars from a Humboldt County retail nursery in northern California. We genotyped isolates and placed them into clonal lineages using microsatellite markers developed for P. ramorum (3,4). Genomic DNA was extracted from mycelia with the FastDNA SPIN kit (Q-Biogene, Morgan, Irvine, CA). Primers used were PrMS6, Pr9C3, PrMS39, PrMS43a, PrMS43b, and PrMS45 (3) and 18, 64, and 82 (4). We sized fluorescently labeled amplicons using capillary electrophoresis (3100 Avant Genetic Analyzer, Applied Biosystems, Foster City, CA). Isolate genotypes were compared with control isolates of known clonal lineage, including BBA9/95 (EU1), Pr102 (NA1), and WSDA3765 (NA2). Three of four isolates belonged to genotype EU1. The fourth isolate, obtained from O. fragrans, belonged to genotype NA1. We repeated genotyping on independent genomic DNA extractions and obtained identical results. Two EU1 isolates and the single NA1 isolate were tested for mating type (1) and found to be of A1, A1, and A2 mating type, respectively. The coexistence of A1 and A2 mating types in the same retail nursery suggests the potential for sexual reproduction, as is the case in P. infestans where clonal and sexual populations exist (2), although to date, sexual reproduction in nature has not been documented in P. ramorum. The California retail nursery infestation highlights the risks associated with the unintentional transport of host nursery stock infested with P. ramorum. References: (1) C. M. Brasier and S. Kirk. Mycol. Res. 108:823, 2004. (2) N. J. Grünwald and W. G. Flier. Ann. Rev. Phytopathol. 43:171, 2005. (3) K. Ivors et al. Mol. Ecol. 15:1493, 2006. (4) S. Prospero et al. Mol. Ecol. 16:2958, 2007.

Effect of portacaval anastomosis on the activities of hepatic enzymes related to cholesterol and bile acid metabolism in rats.

1. The effect of a portacaval anastomosis on the activities of hepatic enzymes related to cholesterol metabolism was investigated in rats. 2. Portacaval anastomosis led to a fall in body weight and liver weight/body weight ratio, and to a rise in the activities of hydroxymethylglutaryl-CoA reductase and cholesterol 7alpha-hydroxylase per g of liver. The net effect was to maintain a normal activity of both enzymes per 100 g of rat. Diurnal rhythm in the activities of both enzymes was maintained after portacaval anastomosis. 3. The rate of excretion of total bile acids, per 100 g of rat, in bile fistula rats was not significantly decreased by portacaval anastomosis.

The effect of portacaval anastomosis on plasma lipoprotein metabolism in rats.

1. The effect of portacaval anastomosis on the metabolism of plasma lipoproteins was investigated in rats. 2. When compared with sham-operated pair-fed controls, plasma high density lipoprotein cholesterol concentration was decreased, plasma low density lipoprotein cholesterol concentration was increased and plasma total cholesterol concentration was unchanged in the portacaval anastomosis rats. Maximal incorporation of [14C]leucine into the total circulating mass of protein was decreased in the very low density lipoprotein and high density lipoprotein fractions and, possibly, in the low density lipoprotein fraction, but there was no change in maximal incorporation into albumin. It is concluded that portacaval anastomosis diminishes the rate of synthesis of high density lipoprotein and very low density lipoprotein proteins and, possibly, of low density lipoprotein proteins.

Effect of diabetes and its control on insulin-like growth factors in the young subject with type I diabetes.

The influence of diabetes and its control on circulating levels of growth hormone and growth hormone-dependent, insulin-like growth factors (IGF) remains controversial. In the present study, the effect of a 1-wk period of intensive insulin therapy on growth hormone and IGF I and II has been determined in 19 young (age 13-22 yr), insulin-dependent (type I) subjects with diabetes mellitus. IGF I was low during conventional insulin therapy (198 +/- 20 versus 438 +/- 38 ng/ml in nondiabetic subjects, P less than 0.001), and rose within the week of intensified treatment (to 255 +/- 15 ng/ml, P less than 0.005), concomitant with a reduction in plasma glucose from 233 +/- 16 to 110 +/- 5 mg/dl. IGF I rose despite a significant fall in mean 24-h growth hormone levels from 14.1 +/- 2.2 to 9.0 +/- 1.2 ng/ml (P less than 0.02). The mean IGF II value for the diabetic subjects (504 +/- 39 ng/ml) was not significantly different from that of the nondiabetic control group (506 +/- 30 ng/ml, P greater than 0.3) and was not altered by intensified therapy. However, four individual patients with very low IGF I also had depressed IGF II (248 +/- 16 ng/ml), which was corrected (to 377 +/- 35 ng/ml) with improved metabolic control. These data suggest that elevated growth hormone levels in poorly controlled diabetes are ineffective in IGF I generation and that this defect is at least partially corrected by acute improvement in control. The rise in IGF I levels accompanying intensive insulin treatment may suppress the excessive secretion of growth hormone.(ABSTRACT TRUNCATED AT 250 WORDS)

Suppression of the growth hormone (GH) response to clonidine and GH-releasing hormone by exogenous GH.

GH release in response to clonidine and human GH-releasing hormone-(1-44) (hGHRH-44) was assessed in 11 boys (aged 7-14 yr) with short stature, who had normal GH secretion. The response to these 2 provocative stimuli was repeated after, respectively, 2 and 3 days of treatment with human GH (0.1 U/kg, im). Exogenous GH significantly blunted the response to both clonidine [the mean 2-h integrated serum GH concentration falling from 1050 +/- 350 (+/- SEM) to 749 +/- 297 ng/ml X min; P = 0.03] and hGHRH-44, the 2-h integrated GH concentration falling from 1553 +/- 358 to 547 +/- 202 ng/ml X min; (P = 0.03). Plasma insulin-like growth factor (IGF-II) concentrations did not change after GH administration. In contrast, plasma IGF-I (somatomedin-C) concentrations increased from 97 +/- 16 ng/ml before administration of GH to 142 +/- 32 ng/ml (P = 0.05) after two days and 149 +/- 23 ng/ml (P less than 0.01) after the third treatment day. However, no correlation was found between the changes in response to clonidine or hGHRH-44 and changes in circulating levels of IGF-I. Our data confirm the existence of GH-dependent feedback inhibition of GH release during childhood and suggest that this inhibition operates, at least in part, at the level of the pituitary. While participation of the IGFs/somatomedins in this feedback loop cannot be excluded, the inhibitory effects of exogenous GH do not depend directly on circulating plasma IGF-I or IGF-II levels.

Ontogeny of leukocyte populations in the ileal Peyer's patch of sheep.

Enzyme- and immunohistochemical methods were used to characterize the leukocyte populations present in the ileal Peyer's patches of sheep foetuses between 68 and 135 d of gestation and particularly in the period around 100 d of gestation, when active lymphopoiesis begins. A wide variety of leukocytes including IgM+, CD5+, CD4+, CD8+ cells, and MgATPase+ dendritic cells were present at an early stage. Groups of IgM+ cells were seen immediately beneath the epithelium as early as 70 d of gestation. Conventional morphometric and computer-assisted morphometric techniques were used to confirm the significant expansion of these cell populations from 90 d of gestation. IgM+ and CD5+ cells were responsible for the vast majority of the increase in cell numbers. It was concluded that a diverse leukocyte population was present at the initiation of active lymphopoiesis in the ileal PP of the sheep foetus and that all members of this population were associated with the emergence of the dome/follicle primordia from which the B-cell follicle develops.

Compartments within the lymph node cortex of calves and adult cattle differ in the distribution of leukocyte populations: an immunohistochemical study using computer-assisted morphometric analysis.

The combination of an immunohistochemical technique and a panel of monoclonal antibodies was used to investigate the presence of leukocyte populations in the distal jejunal lymph node of 3-4 week old calves and adult cattle. The application of computer-assisted morphometric analysis enabled information to be obtained on the distribution of leukocyte populations in lymphoid compartments of the lymph node cortex. Semi-quantitative estimates of the areas of staining in histological sections showed that calves possessed significantly fewer B-cells and CD4+ cells in the outer cortex and significantly fewer T-cells (CD4+, CD8+ and gamma delta T-cells) in the deep cortex. These findings were interpreted to be a possible consequence of immunosuppression resulting from the passive transfer of maternal immunity in colostrum. The presence of some B-cell follicles in the region defined as the deep cortex suggested the on-going differentiation of this predominantly T-cell compartment. The larger presence of interdigitating cells (IDC) in the deep cortex of calves than adults was suggested by significantly larger CD1+ populations and it was argued that this could be the result of the confrontation with exogenous antigen faced by calves in early postnatal life. Antigen presenting populations, pan MHC II+ and MHC II DQ+ populations, were increased in all compartments of calf lymph nodes but were not significantly different from the populations in adult lymph nodes. Variance component analysis of the data generated in the present study showed that the image analysis technique was an effective and statistically powerful approach to investigate leukocyte populations within the specific microenvironments of the lymph node.

Increased gammadelta T-cell populations in draining lymph nodes of lambs during the elicitation phase of dinitrochlorobenzene-induced contact hypersensitivity.

Ten lambs were sensitised with the hapten DNCB in an acetone/olive oil vehicle. The hapten/vehicle solution was applied onto the skin on the shaved ventral surface of the right ear. Two weeks later these lambs were rechallenged with the DNCB/vehicle solution. Simultaneously, ten non-sensitised lambs were treated with vehicle only, serving as vehicle controls. The 20 lambs were slaughtered 48 h after challenge/vehicle treatment, along with ten untreated animals serving as normal controls. Specimens of draining lymph nodes were collected from the 30 animals. All lambs were between 149 and 187 days old. Lymph node cryosections were stained for several leukocyte markers using monoclonal antibodies with the ABC immunohistochemical method. The stained sections were subsequently assessed in three different cortical compartments in each section, using an image analysis system. The resulting measurements from the three groups were compared. A marked increase of gammadelta T cells was detected in the DNCB group. The number of CD4+ T helper cells was decreased in the DNCB group compared with the normal control group, but not with the vehicle control group. No differences were revealed for CD8+ T cytotoxic cells or B cells. These findings were interpreted to be the consequences of possible downregulatory mechanisms protecting the lymphoid tissue from hypersensitivity. The prominence of gammadelta T-cells could indicate that these cells are involved in downregulation.

Gastropathies in the Lundehund. I. Gastritis and gastric neoplasia associated with intestinal lymphangiectasia.

The results from gross and microscopic examination of the stomachs of Lundehunds and examination of stomachs of control dogs from other breeds were compared. Over a 13-year period, 12 of 14 autopsied Lundehunds have been diagnosed as having intestinal lymphangiectasia. In the present study, histological examination revealed gastritis as manifested by an increase in the number of mononuclear cells infiltrating the lamina propria in all the Lundehunds. The inflammation was chronic and restricted to the fundic and body regions, except in one Lundehund where antral gastritis was also present. Computer-assisted morphometric analysis was used to quantify the increased number of mononuclear cells. Atrophy of mucosal fundic glands was prominent in most Lundehunds and mucous metaplasia was often present. Conventional morphometry revealed a significant decrease in the height of the gastric mucosa. A relative expansion in area of the basal part of the lamina propria in Lundehunds with chronic atrophic gastritis corresponded to the observed increase in mononuclear cells and stromal elements. Primary gastric carcinoma with neoplastic cells infiltrating layers of the stomach wall was found in four Lundehunds. The high incidence of gastric carcinoma and the consistent presence of gastritis in Lundehunds suffering from intestinal lymphangiectasia suggest that these changes represent features of a single pathogenetic process.

Gastropathies in the Lundehund. II. A study of mucin profiles.

The mucin profiles of the gastric mucosa in Lundehunds suffering from intestinal lymphangiectasia were examined and compared to the mucin profiles in control dogs from other breeds. A previous study performed on this material had shown that all examined Lundehunds had gastritis and about 30% had gastric carcinoma. Neutral and acid mucins were identified using the periodic acid-Schiff (PAS) and Alcian blue (pH 2.5) periodic acid-Schiff (AB-PAS) methods. The acid mucins were divided into sialomucins and sulfomucins based on their reaction with high-iron diamine Alcian blue, pH 2.5 (HID-AB). In Lundehunds with chronic atrophic gastritis in the fundic and body regions the surface and foveolar epithelium showed a predominantly normal mucin profile although some Lundehunds had a reduced mucin content. The mucous neck cells extended from below the gastric foveolae towards the muscularis mucosae. Morphometric examination showed that the abnormal presence of mucous neck cells occupied 41% of the height of the gastric mucosa in Lundehunds compared to only 19% in the control dogs (p < 0.05). Of the four Lundehunds with gastric carcinoma, two possessed neoplastic cells that contained minimal or no mucins. The amount and type of mucin in the neoplastic cells of the remaining two Lundehunds varied both between individuals and within a neoplasm. This study shows that the abnormal presence of mucous neck cells and the associated pseudopyloric metaplasia comprised the predominant changes in the gastric mucin profiles of Lundehunds.

Distribution of viral antigen in uterus, placenta and foetus of cattle persistently infected with bovine virus diarrhoea virus.

The tissue distribution and cellular localisation of bovine virus diarrhoea virus (BVDV) was investigated in the uterus, placentomes, intercotyledonary foetal membranes and foetal organs of three persistently infected (PI) pregnant heifers. The uterus and ovaries of a non-pregnant PI heifer were also included in the study. Cryostat sections were examined using immunohistochemical techniques and monoclonal antibodies against BVDV. A double immunofluorescence technique was used to identify BVDV positive cells that also showed staining for either the leukocyte common antigen CD45 or the cytoskeletal filament vimentin. BVDV antigen was detected in all the organs examined, and was present in both epithelial and non-epithelial cells. In all organs many of the virus-positive cells also showed reactivity for vimentin. In the foetal liver and spleen a small, scattered population of virus-positive cells showed reactivity for CD45. A few cells showed reactivity both for BVDV antigen and for CD45 in the placentomes and intercotyledonary foetal membranes. In contrast to earlier reports, only scattered cells in the foetal part of the placentomes, the cotyledons, showed reactivity for BVDV antigen. However, in the chorion of the intercotyledonary foetal membranes, a larger proportion of the trophoblast cells showed reactivity for BVDV, especially the binuclear trophoblast cells. In the uterus, pregnancy appeared to favour virus replication, as the section from the pregnant heifers showed much stronger staining and a higher proportion of viral antigen-positive cells than sections from the non-pregnant PI heifer.

Distribution of MHC-II and CD1 molecules in the skin of lambs and changes during experimentally-induced contact hypersensitivity.

The presentation of antigen to specific T-cell populations is a crucial event during the elicitation phase of contact hypersensitivity (CHS). Significant changes in CD4(+) T-cell and gammadelta T-cell populations occur in the skin of sheep 48h after re-exposure to dinitrochlorobenzene but the expression of antigen presentation molecules such as MHC-II and CD1 at this stage of the hypersensitivity response has not been investigated. In the present study, a panel of monoclonal antibodies recognising CD1 and MHC-II subtypes was used in combination with computer assisted morphometric analysis to estimate the distribution of antigen presentation molecules in the superficial and deep dermis of the ears of lambs during the elicitation phase of CHS. The MHC-II molecules showed predominantly a perivascular and peri-appendageal distribution in the dermis and there were scattered MHC-II(+) cells in the basal and suprabasal layers of the epidermis. The CD1w2(+) (CD1b-like) molecules were present on distinct cells that were scattered evenly through the dermis, whereas CD1w3(+) (CD1c-like) molecules were almost exclusively detected on or in close association with the vascular endothelium. There was a significant increase in the presence of MHC-DQ(+) cells in the superficial dermis of dinitrochlorobenzene-treated animals compared with both an untreated control group and a vehicle-treated control group. However, MHC-DQ/DR(+) and CD1w3(+) cells only showed a significant increase compared with the vehicle-treated control group. The present study shows that the distribution of molecules involved in antigen presentation to CD4(+) T-cells and gammadelta T-cells changes during the elicitation phase of CHS in sheep, and suggests a role for MHC-DQ molecules on antigen presenting cells. However, the changes in distribution and expression of MHC-II and CD1 subtypes argue against a prominent role for a CD1-dependent pathway for T-cell recognition in the clinical cutaneous hypersensitivity response in sheep. Based on the expression of MHC-II molecules and CD1c molecules, we also suggest a potential role for endothelial cells in antigen presentation during the clinical dermatitis reaction.

Factor XIIIa positive dendritic cells are a major accessory cell population in the elicitation phase of DNCB-induced contact hypersensitivity.

The phenotypes and distribution of accessory cells in the ear skin of lambs during the elicitation phase of dinitrochlorobenzene (DNCB)-induced contact hypersensitivity (CHS) were examined using indirect immunoperoxidase histochemistry (ABC method), and a panel of antibodies. Thirty lambs, between 21 and 26 weeks of age, were divided into groups of 10. The shaved right ear of one group was treated with DNCB. Two weeks later this group was challenged with DNCB. One group was treated with the vehicle alone and the remaining group was left untreated. The lambs were slaughtered 48 h after challenge, and tissue specimens were collected from the ears of the three groups. Factor XIIIa+ (FXIIIa+) cells were prominent in the superficial dermis and showed predominantly a perivascular and subepidermal distribution. The other markers were less prominent, and whereas CD1+ cells and CD68+ cells showed a reaction pattern similar to the FXIIIa+ cells, CD14+ cells were found scattered predominantly in the deep dermis. There appeared to be an increase in FXIIIa+ cells, CD1+ cells, and CD68+ cells in the dermis of the DNCB-treated lambs 48 h after challenge. Only CD1+ cells were detected in epidermis of normal controls, and these cells appeared to be decreased in number in the two treated groups. Computer-assisted morphometric analysis was used to estimate the relative presence of the accessory cell subpopulations in the superficial and deep dermis and the entire dermis. A statistical analysis of the relative area of immunostaining showed a significantly increased presence of FXIIIa+ cells and CD68+ cells in the dermis of the DNCB-treated lambs 48 h after challenge. Interestingly, FXIIIa+ cells and CD68+ cells were also significantly increased in the vehicle treated group compared with untreated controls. We found no significant difference in the presence of CD1+ cells or CD14+ cells in the DNCB treated group compared with the controls. The study showed that FXIIIa+ DDC are the major accessory cell population in normal ear skin of lambs and the major responsive population during the elicitation phase of CHS. The lack of response in the CD1+ cell population suggests a less prominent role for the LC-related DC in the skin during the elicitation phase.

Accessory cell populations in draining lymph nodes of lambs in the elicitation phase of DNCB-induced contact hypersensitivity.

The effect of experimentally induced contact hypersensitivity on accessory cell populations in draining lymph nodes of lambs was studied. Previous studies of draining lymph nodes of lambs during the elicitation phase of CHS have shown that there are significant changes in T-cell subpopulations, particularly CD4(+) cells and gamma delta T-cells, but the behaviour of accessory (antigen presenting) cell populations was not investigated. The immunohistochemical presence of accessory cell populations was determined using markers for CD68, Pan MHCII, MHCII DQ, MHCII DR, OvCD1w1 (putative human CD1a/c-like) and OvCD1w2 (human CD1b-like). Ten lambs were sensitised, and 14 days later re-challenged, by applying the hapten di-nitro-chloro-benzene (DNCB) together with an acetone and olive oil (AOO) vehicle, onto the skin. Cryosections of the draining lymph nodes were stained immunohistochemically for the accessory cell markers. Using an image analysis system, the areas of staining in the lymph nodes from the challenged animals were compared with measurements in control animals. A significant increase in staining for CD68(+) cells was detected in the cortex of the DNCB-treated group (p=0.003). A significant increase in staining for the Pan MHCII marker was also observed in the DNCB group (p=0. 013). These results show that MHCII(+) cells and CD68(+) cells constitute a prominent cell population in the cortex of the regional lymph nodes of lambs in the late elicitation phase of DNCB-induced contact hypersensitivity.

Lymphoid follicles in the gastric mucosa of dogs. Distribution and lymphocyte phenotypes.

The occurrence and distribution of lymphoid follicles within the stomachs of 36 dogs that did not have macroscopic gastric lesions are presented. The dogs ranged in age from less than 1 year to over 13 years. The number of follicles varied between the different regions of the stomach, being most numerous (15.6 follicles cm-2) and uniform in size (about 1 mm in diameter) in the fundus. The number and size of follicles in the antrum varied widely between dogs. Age-related changes in the distribution of follicles were not found following simple linear regression analysis. The phenotypes of lymphocytes in gastric lymphoid follicles of nine dogs aged from less than 1 year to 5 years were determined using monoclonal antibodies specific for canine leucocyte antigens and an indirect immunoperoxidase technique. The follicles had an organized distribution of lymphocytes subsets in that a predominantly B cell area contained some CD4+ cells and very few CD8+ cells and was adjacent to an area containing mostly T cells. Computer-assisted morphometric analysis was used to quantify the overall presence of the various lymphocyte subpopulations. Follicles in the fundus and body regions possessed similar percentages of lymphocytes averaging 42%, 22% and 3% of the area occupied by B cells, CD4+ and CD8+ cells, respectively. It is concluded that lymphoid follicles are a normal constituent of the canine gastric mucosa and possess a lymphocyte composition similar to that reported by others for solitary intestinal follicles.

Spleen and kidney of Atlantic salmon (Salmo salar L.) show histochemical changes early in the course of experimentally induced infectious salmon anaemia (ISA).

Infectious salmon anaemia (ISA) is a disease of farmed Atlantic salmon (Salmo salar L.) in Norway that affects both erythrocytic and leucocytic cells. Both cell types are possible target cells for the aetiological ISA agent, which is probably a virus. In the present study the distribution and phenotype of leucocyte populations in the spleen and head kidney of Atlantic salmon that were developing ISA have been examined. Frozen tissues were collected from fish at various times after inoculation with ISA-infective material. Immune and enzyme histochemical techniques were used to characterise the response of leucocyte populations. Acid phosphatase positive macrophages predominantly in the red pulp of the spleen appeared to have engulfed erythrocytes at day 4 after infection. Evidence of degradation products of phagocytosed erythrocytes was present in macrophages in red pulp of the spleen at day 7 after infection, in addition to the usual site of erythrophagocytosis in melanomacrophage accumulations. Signs of erythrophagocytosis were not found in the head or body portions of the kidney. The activation of macrophages in the spleen at day 7 was suggested by decreased reactivity for the enzyme 5' nucleotidase. From day 7, clusters of immunoglobulin positive (Ig +) cells were present in the head kidney, while from day 11, the ellipsoids of the spleen showed reactivity for Ig and complement factor C3. These observations are discussed in relation to early immunoglobulin production and possible immune complex trapping. The present results suggest that the leucocyte populations in Atlantic salmon respond to ISA infection through macrophage activation and the initiation of an immune response.

Subclinical paratuberculosis in goats following experimental infection. An immunological and microbiological study.

An experimental oral infection of goats with a caprine isolate of Mycobacterium a. subsp. paratuberculosis was used to investigate immunological and bacteriological events during the subclinical phase of infection. Seven goats at 5-8 weeks of age were given a bacterial suspension in milk-replacement three times weekly for 9 weeks. Six animals were kept as controls. Cellular recall responses against M. a. paratuberculosis were analysed by means of a lymphocyte proliferation test, an IFN-gamma assay and an IL-2 receptor assay. All inoculated animals had detectable CMI responses from 9 weeks post-inoculation and through the 2 years of study, although the responses were highest during the first year. Antibodies against M. a. paratuberculosis could be detected from weeks 15-20 in four of the seven animals, and one additional animal became antibody positive at week 35, while two inoculated animals did not produce significant antibody titres during the experiment. At about 1-year post-inoculation, two animals became faecal shedders, while two others started to excrete bacteria into faeces about 2 years post-inoculation. The appearance of M. a. paratuberculosis in faeces was not associated with a decline in cellular responses as far as could be assessed using the current methods for measuring CMI. Pathological lesions due to M. a. paratuberculosis infection and presence of bacteria were recorded in the intestine and/or mesenteric lymph nodes of five animals while lymph node changes suggestive of paratuberculosis were observed in one animal. Only the two animals with no signs of an active infection at necropsy showed a considerable decline in the cellular parameters during the last year of the study, particularly in the IFN-gamma assay. The two animals with the highest levels of M. a. paratuberculosis responsive CD8+ lymphocytes in the circulation about 1-year post-inoculation had no detectable lesions in the distal ileum and colon at necropsy, while high numbers of gammadelta T-cells responsive to M. a. paratuberculosis in the circulation were associated with disseminated lesions in the distal ileum and colon.

Cellular composition of utero-ovarian lymph during the exogenous gonadotrophin-stimulated oestrous cycle in sheep.

Utero-ovarian peripheral lymph was collected for extended periods from sheep that had received exogenous gonadotrophins. High lymph flow rates and progesterone outputs, over 13 mL h-1 and 65 nmol h-1 respectively, were observed during the subsequent oestrous cycles. A positive correlation was found between the maximum lymph flow rate and the number of corpora lutea in the draining ovary. At periods approximating the beginning and end of the luteal phase of the oestrous cycle there were increases in the output of red blood cells and, towards the end of the luteal phase, eosinophils constituted over 10% of the white blood cells in the lymph. Lymph provides a dynamic record of the changing internal environment of the ovary and as such may yield useful information on intraovarian control mechanisms in sheep.